Safety Risks: Minimal

The air pumps make a loud noise and lithium ion batteries require care when handling. Roses have sharp prickles, wear gloves to protect from scratches. Hearing protection is recommended. There may be a risk of electric shock in rainy conditions. Wear appropriate protection from the sun: sunscreen and water are recommended.

Explanation of rationale behind experimental design:

For this experiment, we will be collecting headspace volatiles with Solid Phase Microextraction (SPME), a method which adsorbs the volatile organic compounds (VOCs) surrounding part of a plant to the SPME fiber. The adsorbed chemicals can then be desorbed with paired Gas Chromatography - Mass Spectrometery (GC-MS) equipment to identify the collected chemistry. SPME extraction is a flexible method which can collect the VOCs from just about anything you can get an container around. For our experiments, we rely on heat-treated oven bags, which are relatively chemically inert after cooking them in an oven at 50°C for a day or so.

The purpose of these experiments is to collect the volatiles emanating from roses under various conditions, including:

Goal of study:

By sampling rose VOCs with SPME under these different conditions, we hope to find quantitative and/or qualitative differences in VOC compounds between rose treatments.

Hypotheses:

Our hypothesis formed from preliminary research where we observed that A. swirskii were preferentially attracted the RRD-infected roses. We think this might be due to differences in the VOCs released from uninfected vs. infected roses. Collecting and comparing chemistry between different roses should allow us to see any qualitative and/or quantitative differences in VOCs released between these groups. This data will give us insight into some of the metabolic changes which may have occurred between different rose groups. Furthermore, our data should indicate which chemicals are worth testing for attractiveness to A. swirskii in olfactometer trials, which can give us insight into which specific VOCs A. swirskii may have been responding to from the RRD-infected roses.

Materials:

Clean air system:
  • 3 high volume air inflators (24v, 25 CFM) w/ nozzle attachments
  • 6-8 lithium ion (24v, 4.0 Ah) power tool batteries (charged)
  • 2 variable flow meters with PTFE tubing
  • 2 In line Hepa Sanitary Air Filters for 1/4 in tubing (attached to flow meter)
  • 2 inline water filter carbon blocks, one micron filtration
Headspace volatile collection equipment:
  • GoBeGreen Original Garden Bags - Nylon - One per sample + Backups (precooked in oven at 50°C for 24 hours or more)
  • Zip ties - 6 + per each sample, bring a large number
  • Heavy duty shears (for cutting zip ties and holes in the bags)
  • Bypass pruners (optional, for removing dead canes, allowing better access to rose flowers)
  • Blue mini inline filters (for outflow)
Solid Phase Microextraction (SMPE)
  • Manual SPME holder
  • SPME fiber (Gray Fiber) - 24 Ga Divinylbenzene/Carboxen/Polydimethylsiloxane (DVB/CAR/PDMS)
  • 2 Double Burette clamps
  • 2 Aluminum Rod (3 ft, 1/4 in)
Eqiupment for Internal Standard:
  • Nonyl acetate (keep cool)
  • 1 Drummond Scientific Aspirator, Captrol III
  • 5 ul glass calibrated micropipettes

Methods:

To begin, select a cluster of flowers from a single rose cane, and gently cover it with an oven bag. Seal the end of the oven bag over the base of the rose cane with a zip tie so that the air is trapped around the

Experimental Site:

Notes and Figures:

Assembled Field Extractor:

Figure 1: Assembled clean air delivery system for in-field extraction of rose headspace volatiles via SPME

Burette holder

Figure 2: The burette holder can be adjusted to different heights to accommadate the SPME holder in a variety of positions. Note the placement of the hoses and the blue inline filter.

Components of SPME holder:

Figure 3: The SPME fibers are really fragile: please protect the needles from bending and make sure that the fibers do not come into contact with the plant during use.

Risks of overheating:

Figure 4: The air pumps will overheat in direct sunlight, especially when ambient temperatures are over 32°C (90°F). To prevent damaging the equipment, try to keep inflators insulated from heat, and in the shade if possible. Take care to avoid obstructing the intake vents. When the air pumps overheat, it melts the internal fans and creates a situation where the motor melts through the internal wiring, completing circuit and preventing the power switch from functioning while the battery is still attached. If you are not able to turn off the air pump with the toggle switch (without removing the battery), then the air pumps are permanently damaged, and should be replaced immediately. Also, you will need to remove any particulates of melted fan matter from the carbon block filters (just shake it out).

last modified: 02 July 2021